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Shigella Effector OspB Activates mTORC1 in a Manner That Depends on IQGAP1 and Promotes Cell Proliferation.

Lu R, Herrera BB, Eshleman HD, Fu Y, Bloom A, Li Z, Sacks DB, Goldberg MB - PLoS Pathog. (2015)

Bottom Line: We show that the effect on the area of bacterial spread is due to OspB triggering increased cell proliferation at the periphery of infected foci, thereby replacing some of the cells that die within infected foci and restricting the area of bacterial spread.OspB activation of mTORC1, and its effects on cell proliferation and bacterial spread, depends on IQGAP1.Our results identify OspB as a regulator of mTORC1 and mTORC1-dependent cell proliferation early during S. flexneri infection and establish a role for IQGAP1 in mTORC1 signaling.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Cambridge, Massachusetts, United States of America; Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT
The intracellular bacterial pathogen Shigella infects and spreads through the human intestinal epithelium. Effector proteins delivered by Shigella into cells promote infection by modulating diverse host functions. We demonstrate that the effector protein OspB interacts directly with the scaffolding protein IQGAP1, and that the absence of either OspB or IQGAP1 during infection leads to larger areas of S. flexneri spread through cell monolayers. We show that the effect on the area of bacterial spread is due to OspB triggering increased cell proliferation at the periphery of infected foci, thereby replacing some of the cells that die within infected foci and restricting the area of bacterial spread. We demonstrate that OspB enhancement of cell proliferation results from activation of mTORC1, a master regulator of cell growth, and is blocked by the mTORC1-specific inhibitor rapamycin. OspB activation of mTORC1, and its effects on cell proliferation and bacterial spread, depends on IQGAP1. Our results identify OspB as a regulator of mTORC1 and mTORC1-dependent cell proliferation early during S. flexneri infection and establish a role for IQGAP1 in mTORC1 signaling. They also raise the possibility that IQGAP1 serves as a scaffold for the assembly of an OspB-mTORC1 signaling complex.

No MeSH data available.


Related in: MedlinePlus

OspB activates mTOR during S. flexneri infection.(A-B) Phosphorylation of S6K during infection of IQGAP1-/- and IQGAP1+/+ MEFs with non-invasive, wild type (WT), ΔospB (ospB), or ΔospB complemented with p-ospB (ospB p-OspB) S. flexneri (A), and in the presence or absence of 10 nM rapamycin (B). Apparent MW are in Kd. (C) Area of plaques formed in monolayers by wild type or ΔospB S. flexneri in the presence or absence of 10 nM rapamycin. Data represent mean ± S.D. of three or more independent experiments. *, p < 0.05, Student’s two-tailed t test.
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ppat.1005200.g005: OspB activates mTOR during S. flexneri infection.(A-B) Phosphorylation of S6K during infection of IQGAP1-/- and IQGAP1+/+ MEFs with non-invasive, wild type (WT), ΔospB (ospB), or ΔospB complemented with p-ospB (ospB p-OspB) S. flexneri (A), and in the presence or absence of 10 nM rapamycin (B). Apparent MW are in Kd. (C) Area of plaques formed in monolayers by wild type or ΔospB S. flexneri in the presence or absence of 10 nM rapamycin. Data represent mean ± S.D. of three or more independent experiments. *, p < 0.05, Student’s two-tailed t test.

Mentions: To test the relevance of OspB activation of mTOR during S. flexneri infection, we compared S6K phosphorylation in cells infected with an ospB mutant to that in cells infected with WT S. flexneri or the complemented ospB mutant. At 1 hr of infection of IQGAP1+/+ MEFs, WT or ospB complemented S. flexneri induced more phosphorylation of S6K than the ospB mutant (Fig 5A), indicating that OspB delivered by S. flexneri activates mTOR early during infection. Activation of mTOR during infection depended on IQGAP1, since S6K phosphorylation remained at baseline in IQGAP1-/- MEFs. For cells that were exposed to the non-invasive S. flexneri mutant BS103, S6K phosphorylation was lower than baseline, suggesting that the presence of extracellular bacteria might suppress mTOR activation, perhaps by reducing the concentration of amino acids in the extracellular media [28]. Treatment of cells with rapamycin inhibited OspB+ S. flexneri-induced phosphorylation of S6K (Fig 5B), indicating that S6K phosphorylation was mediated by mTOR kinase. Rapamycin had no effect on bacterial growth in vitro (S6A Fig).


Shigella Effector OspB Activates mTORC1 in a Manner That Depends on IQGAP1 and Promotes Cell Proliferation.

Lu R, Herrera BB, Eshleman HD, Fu Y, Bloom A, Li Z, Sacks DB, Goldberg MB - PLoS Pathog. (2015)

OspB activates mTOR during S. flexneri infection.(A-B) Phosphorylation of S6K during infection of IQGAP1-/- and IQGAP1+/+ MEFs with non-invasive, wild type (WT), ΔospB (ospB), or ΔospB complemented with p-ospB (ospB p-OspB) S. flexneri (A), and in the presence or absence of 10 nM rapamycin (B). Apparent MW are in Kd. (C) Area of plaques formed in monolayers by wild type or ΔospB S. flexneri in the presence or absence of 10 nM rapamycin. Data represent mean ± S.D. of three or more independent experiments. *, p < 0.05, Student’s two-tailed t test.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4608727&req=5

ppat.1005200.g005: OspB activates mTOR during S. flexneri infection.(A-B) Phosphorylation of S6K during infection of IQGAP1-/- and IQGAP1+/+ MEFs with non-invasive, wild type (WT), ΔospB (ospB), or ΔospB complemented with p-ospB (ospB p-OspB) S. flexneri (A), and in the presence or absence of 10 nM rapamycin (B). Apparent MW are in Kd. (C) Area of plaques formed in monolayers by wild type or ΔospB S. flexneri in the presence or absence of 10 nM rapamycin. Data represent mean ± S.D. of three or more independent experiments. *, p < 0.05, Student’s two-tailed t test.
Mentions: To test the relevance of OspB activation of mTOR during S. flexneri infection, we compared S6K phosphorylation in cells infected with an ospB mutant to that in cells infected with WT S. flexneri or the complemented ospB mutant. At 1 hr of infection of IQGAP1+/+ MEFs, WT or ospB complemented S. flexneri induced more phosphorylation of S6K than the ospB mutant (Fig 5A), indicating that OspB delivered by S. flexneri activates mTOR early during infection. Activation of mTOR during infection depended on IQGAP1, since S6K phosphorylation remained at baseline in IQGAP1-/- MEFs. For cells that were exposed to the non-invasive S. flexneri mutant BS103, S6K phosphorylation was lower than baseline, suggesting that the presence of extracellular bacteria might suppress mTOR activation, perhaps by reducing the concentration of amino acids in the extracellular media [28]. Treatment of cells with rapamycin inhibited OspB+ S. flexneri-induced phosphorylation of S6K (Fig 5B), indicating that S6K phosphorylation was mediated by mTOR kinase. Rapamycin had no effect on bacterial growth in vitro (S6A Fig).

Bottom Line: We show that the effect on the area of bacterial spread is due to OspB triggering increased cell proliferation at the periphery of infected foci, thereby replacing some of the cells that die within infected foci and restricting the area of bacterial spread.OspB activation of mTORC1, and its effects on cell proliferation and bacterial spread, depends on IQGAP1.Our results identify OspB as a regulator of mTORC1 and mTORC1-dependent cell proliferation early during S. flexneri infection and establish a role for IQGAP1 in mTORC1 signaling.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Cambridge, Massachusetts, United States of America; Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT
The intracellular bacterial pathogen Shigella infects and spreads through the human intestinal epithelium. Effector proteins delivered by Shigella into cells promote infection by modulating diverse host functions. We demonstrate that the effector protein OspB interacts directly with the scaffolding protein IQGAP1, and that the absence of either OspB or IQGAP1 during infection leads to larger areas of S. flexneri spread through cell monolayers. We show that the effect on the area of bacterial spread is due to OspB triggering increased cell proliferation at the periphery of infected foci, thereby replacing some of the cells that die within infected foci and restricting the area of bacterial spread. We demonstrate that OspB enhancement of cell proliferation results from activation of mTORC1, a master regulator of cell growth, and is blocked by the mTORC1-specific inhibitor rapamycin. OspB activation of mTORC1, and its effects on cell proliferation and bacterial spread, depends on IQGAP1. Our results identify OspB as a regulator of mTORC1 and mTORC1-dependent cell proliferation early during S. flexneri infection and establish a role for IQGAP1 in mTORC1 signaling. They also raise the possibility that IQGAP1 serves as a scaffold for the assembly of an OspB-mTORC1 signaling complex.

No MeSH data available.


Related in: MedlinePlus